| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 3 | John I. Leonard |
A | barrett | castillo | toussaint | louis | 54.32 | John I Leonard Invitational 1 |
| 4 | Pahokee |
A | Jones | Johnson | Washington | White | 50.11 | FHSAA 1A Region 4 |
| 5 | Glades Central |
A | Josinvil | Forbes | Marshall | Butts | 51.17 | Royal Palm Beach Invitational 2026 |
| 6 | Glades Day |
A | Ocanas | Baez | Sturgis | Sisnett | 55.65 | Pahokee Anquan Boldin Invitational 2025 |